The proposed work addresses a pivotal study for decisions on the form of the GeoVax Clade B HIV vaccine (GOVX-B11) to be taken into efficacy trials. GOVX-B11 is a DNA/MVA vaccine that expresses virus-like particles (VLPs) displaying trimeric membrane bound Env. It has been under development for more than 15 years, during which time it has demonstrated unique properties and shown considerable promise in both preclinical and clinical studies. Phase I of this SBIR tested the ability of monovalent gp120 and gp140 boosts to enhance the immunogenicity of GOVX-B11. This Phase II proposal builds on the Phase I results to test a bivalent gp120 boost and to extend preclinical testing to include a challenge study. The Clade B epidemic continues to be a major public health problem in the Americas. Although the US epidemic does not receive the publicity received by the epidemic in Africa, and should theoretically be able to be controlled with drugs, HIV remains a major US health problem. Costs of HIV care and treatment rose 37% from $12 billion in 2009 to just under $17 billion in 2014 - a high and increasing burden for taxpayers (Kaiser Foundation Fact Sheet, June 2014). Financial costs, alone, demonstrate the critical need for a vaccine. Social costs of each HIV infection are also large, including patient time, lost productivity, and physical and emotional distress to patients and their families. Specific Aim 1. To conduct a preclinical trial (GV-M2) testing a bivalent protein boost for GOVX-B11. The proposed bivalent boost consists of a transmitted/founder gp120 (B63521delta11) + a CD4-induced form of Bal gp120 (FLSC). The primary goals of this specific aim are to oversee the conduct of the trial and to collect safet data for a trial that will include the D1D2 domains of CD4 in an immunogen. Specific Aim 2. To obtain immunogenicity data for the bivalent protein boost. The primary goal of this Specific Aim is to test the effect of the bivalent boost on the titer, isotype, specificity, breadth and functio of antibody (Ab) to Env. Particular emphasis will be placed on Ab for the V1V2, V3, and C1 regions of gp120 and the ADCC activity of the elicited Ab, all of which were associated with reduced risk in the partially successful RV144 trial. Specific Aim 3. To determine the effect of the bivalent B63521delta11+FLSC boost on the ability to protect against serial rectal challenges with SHIV-162P3. The primary goals of this Specific Aim are to determine whether the bivalent protein boost increases protection against acquisition and to determine which Ab and T cell responses decrease acquisition and how these responses relate to those that decreased the risk of infection in the RV144 trial.